In vivo photocontrol of orexin receptors with a nanomolar light-regulated analogue of orexin-B.

Orexinergic neurons are critically involved in regulating arousal, wakefulness, and appetite. Their dysfunction has been associated with sleeping disorders, and non-peptide drugs are currently being developed to treat insomnia and narcolepsy. Yet, no light-regulated agents are available to reversibly control their activity. To meet this need, a photoswitchable peptide analogue of the endogenous neuroexcitatory peptide orexin-B was designed, synthesized, and tested in vitro and in vivo. This compound - photorexin - is the first photo-reversible ligand reported for orexin receptors. It allows dynamic control of activity in vitro (including almost the same efficacy as orexin-B, high nanomolar potency, and subtype selectivity to human OX2 receptors) and in vivo in zebrafish larvae by direct application in water. Photorexin induces dose- and light-dependent changes in locomotion and a reduction in the successive induction reflex that is associated with sleep behavior. Molecular dynamics calculations indicate that trans and cis photorexin adopt similar bent conformations and that the only discriminant between their structures and activities is the positioning of the N-terminus. This, in the case of the more active trans isomer, points towards the OX2 N-terminus and extra-cellular loop 2, a region of the receptor known to be involved in ligand binding and recognition consistent with a "message-address" system. Thus, our approach could be extended to several important families of endogenous peptides, such as endothelins, nociceptin, and dynorphins among others, that bind to their cognate receptors through a similar mechanism: a "message" domain involved in receptor activation and signal transduction, and an "address" sequence for receptor occupation and improved binding affinity.

Photoswitchable carbamazepine analogs for non-invasive neuroinhibition in vivo.

A problem of systemic pharmacotherapy is off-target activity, which causes adverse effects. Outstanding examples include neuroinhibitory medications like antiseizure drugs, which are used against epilepsy and neuropathic pain but cause systemic side effects. There is a need of drugs that inhibit nerve signals locally and on-demand without affecting other regions of the body. Photopharmacology aims to address this problem with light-activated drugs and localized illumination in the target organ. Here, we have developed photoswitchable derivatives of the widely prescribed antiseizure drug carbamazepine. For that purpose, we expanded our method of ortho azologization of tricyclic drugs to meta/para and to N-bridged diazocine. Our results validate the concept of ortho cryptoazologs (uniquely exemplified by Carbazopine-1) and bring to light Carbadiazocine (8), which can be photoswitched between 400-590 nm light (using violet LEDs and halogen lamps) and shows good drug-likeness and predicted safety. Both compounds display photoswitchable activity in vitro and in translucent zebrafish larvae. Carbadiazocine (8) also offers in vivo analgesic efficacy (mechanical and thermal stimuli) in a rat model of neuropathic pain and a simple and compelling treatment demonstration with non-invasive illumination.

Correction: Photoswitchable dynasore analogs to control endocytosis with light.

[This corrects the article DOI: 10.1039/D0SC03820B.].

A photoswitchable GABA receptor channel blocker.

BACKGROUND AND PURPOSE: Anion-selective Cys-loop receptors (GABA and glycine receptors) provide the main inhibitory drive in the CNS. Both types of receptor operate via chloride-selective ion channels, though with different kinetics, pharmacological profiles, and localization. Disequilibrium in their function leads to a variety of disorders, which are often treated with allosteric modulators. The few available GABA and glycine receptor channel blockers effectively suppress inhibitory currents in neurons, but their systemic administration is highly toxic. With the aim of developing an efficient light-controllable modulator of GABA receptors, we constructed azobenzene-nitrazepam (Azo-NZ1), which is composed of a nitrazepam moiety merged to an azobenzene photoisomerizable group. EXPERIMENTAL APPROACH: The experiments were carried out on cultured cells expressing Cys-loop receptors of known subunit composition and in brain slices using patch-clamp. Site-directed mutagenesis and molecular modelling approaches were applied to evaluate the mechanism of action of Azo-NZ1. KEY RESULTS: At visible light, being in trans-configuration, Azo-NZ1 blocked heteromeric α1/ß2/γ2 GABAA receptors, ρ2 GABAA (GABAC ), and α2 glycine receptors, whereas switching the compound into cis-state by UV illumination restored the activity. Azo-NZ1 successfully photomodulated GABAergic currents recorded from dentate gyrus neurons. We demonstrated that in trans-configuration, Azo-NZ1 blocks the Cl-selective ion pore of GABA receptors interacting mainly with the 2' level of the TM2 region. CONCLUSIONS AND IMPLICATIONS: Azo-NZ1 is a soluble light-driven Cl-channel blocker, which allows photo-modulation of the activity induced by anion-selective Cys-loop receptors. Azo-NZ1 is able to control GABAergic postsynaptic currents and provides new opportunities to study inhibitory neurotransmission using patterned illumination.